Essentiality Info:
According to the Brenda site, "N-ureido analogue, synthesised by us, had inhibitory activity against the aforementioned de-N-acetylase, presumably via the N-ureido motif."
Is it a Monomer or Multimer?: monomer
Complex of Proteins: N/A
Druggable target: The following info was found on Brenda: "Substrate specificity of the N-acetylglucosaminyl-phosphatidylinositol de-N-acetylase of glycosylphosphatidylinositol membrane anchor biosynthesis in African trypanosomes and human cells." http://www.brenda-enzymes.org/literature.php?e=3.5.1.89&r=649652
EC 3.5.1.89
Disease information
Sleeping sickness, African trypanosomiasis, is a deadly blood disease caused by two variates of Trypanosoma brucei and transmitted by tsetse fly.
In 1–3 weeks after the bite a chancre (a red sore skin lesion) can develop on the bite area. Several weeks or months later Trypanosoma parasites in the blood, spinal and lymphatic fluid (hemolymphatic stage) can cause fever, anemia, etc. As the disease reaches its final stage, the parasites get through the blood-brain barrier entering the brain. This meningoencephalitic stage (inflammation of the central nervous system) could cause blackouts, coma, or death.
Pentamidine isethionate and suramin are usually used for treating the hemolymphatic stage of West and East African Trypanosomiasis, respectively. Melarsoprol is used for late disease with central nervous system involvement.
Tsetse flies are common only in rural Africa, not in urban cities, so this disease is considered an "Old World" disease.
List cost and quantity of substrate reagents, supplier, and catalog #
6-(N-acetyl-alpha-D-glucosaminyl)-1-phophatidyl-1D-myo-inositol: no info found
water: found in lab currently
Structure: N/A
Current inihibitors: "Synthesis of 1-D-6-O-[2-(N-hydroxyaminocarbonyl)amino-2-deoxy-alpha-D-glucopyranosyl]-myo-inositol 1-(n-octadecyl phosphate): a potential metalloenzyme inhibitor of glycosylphosphatidylinositol biosynthesis" http://www.ncbi.nlm.nih.gov/pubmed/18479678
Expression Information: No information was found on expression in other bacteria.
Purification method: E-52 cellulose batch preparation followed by 2' AMP-agarose affinity chromatography
Figure 2: TMpred graph of 2-acetamido-2-deoxy-alpha-D-glucopyranosyl-(1->6)-phosphatidylinositol de-N-acetylase of T. brucei run using amino acid sequence.
CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): N/a
GC% Content for gene (codon optimized): N/a
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to. Primer design results for 'tail' primers (this is just 2 sequences):
NCBI Gene # : 719226
Protein ID: fig|870187.4.peg.785
Organism: Trypanosoma brucei
Etiologic Risk Group: Risk group 2- Parasitic agents
Link to TDR targets page: N/A
Link to Gene database (Patric): https://www.patricbrc.org/portal/portal/patric/Feature?cType=feature&cId=PATRIC.870187.4.AJUL01000009.CDS.733719.734555.rev
Essentiality Info:
According to the Brenda site, "N-ureido analogue, synthesised by us, had inhibitory activity against the aforementioned de-N-acetylase, presumably via the N-ureido motif."
Is it a Monomer or Multimer?: monomer
Complex of Proteins: N/A
Druggable target: The following info was found on Brenda: "Substrate specificity of the N-acetylglucosaminyl-phosphatidylinositol de-N-acetylase of glycosylphosphatidylinositol membrane anchor biosynthesis in African trypanosomes and human cells."
http://www.brenda-enzymes.org/literature.php?e=3.5.1.89&r=649652
EC 3.5.1.89
Disease information
Sleeping sickness, African trypanosomiasis, is a deadly blood disease caused by two variates of Trypanosoma brucei and transmitted by tsetse fly.
In 1–3 weeks after the bite a chancre (a red sore skin lesion) can develop on the bite area. Several weeks or months later Trypanosoma parasites in the blood, spinal and lymphatic fluid (hemolymphatic stage) can cause fever, anemia, etc. As the disease reaches its final stage, the parasites get through the blood-brain barrier entering the brain. This meningoencephalitic stage (inflammation of the central nervous system) could cause blackouts, coma, or death.
Pentamidine isethionate and suramin are usually used for treating the hemolymphatic stage of West and East African Trypanosomiasis, respectively. Melarsoprol is used for late disease with central nervous system involvement.
Tsetse flies are common only in rural Africa, not in urban cities, so this disease is considered an "Old World" disease.
link to Brenda EC page:
http://www.brenda-enzymes.org/enzyme.php?ecno=3.5.1.89
Figure 1: Reaction schematic on Brenda site
Enzyme Assay Info: non-radioactive high-throughput assay
http://www.ncbi.nlm.nih.gov/pubmed/20085486
List cost and quantity of substrate reagents, supplier, and catalog #
6-(N-acetyl-alpha-D-glucosaminyl)-1-phophatidyl-1D-myo-inositol: no info found
water: found in lab currently
Structure: N/A
Current inihibitors: "Synthesis of 1-D-6-O-[2-(N-hydroxyaminocarbonyl)amino-2-deoxy-alpha-D-glucopyranosyl]-myo-inositol 1-(n-octadecyl phosphate): a potential metalloenzyme inhibitor of glycosylphosphatidylinositol biosynthesis"
http://www.ncbi.nlm.nih.gov/pubmed/18479678
Expression Information: No information was found on expression in other bacteria.
Purification method: E-52 cellulose batch preparation followed by 2' AMP-agarose affinity chromatography
Image in Pymol:
Amino Acid Sequence:
0 MCPINASFYT LIVVKLLRES LIHMHGALAF GFVVVFLSFL VLWQRASCVS KIHLVGDVLF
60 VFAHPDDEAM FFSPLLDYVR RHGLNAHFLC LSNGNYSGLG TVREKELVAS AEYFGVNRRS
120 VRVVDHPDLQ DGPDNLWNTE IVQREVLSYL HSVKDIRTVI TFDHRGVSSH ANHVAVYEGV
180 LLAKKNLPPG ILFLSLHTRD LLEKYVGILS TVGYTVGIHR CGGRRNHVIL IPPTSLFTSF
240 SAMRKHKTQL VWFRYLFVWF SSYSYVNEVK ELGVA
Length of Protein: 275
Molecular Weight: 31205 kD
Molar Extinction coefficient of your protein at 280 nm wavelength: 38640
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html):
Figure 2: TMpred graph of 2-acetamido-2-deoxy-alpha-D-glucopyranosyl-(1->6)-phosphatidylinositol de-N-acetylase of T. brucei run using amino acid sequence.
CDS Gene Sequence (paste as text only):
http://www.bioinformatics.org/sms2/rev_trans.html
atgtgcccgattaacgcgagcttttataccctgattgtggtgaaactgctgcgcgaaagc
ctgattcatatgcatggcgcgctggcgtttggctttgtggtggtgtttctgagctttctg
gtgctgtggcagcgcgcgagctgcgtgagcaaaattcatctggtgggcgatgtgctgttt
gtgtttgcgcatccggatgatgaagcgatgttttttagcccgctgctggattatgtgcgc
cgccatggcctgaacgcgcattttctgtgcctgagcaacggcaactatagcggcctgggc
accgtgcgcgaaaaagaactggtggcgagcgcggaatattttggcgtgaaccgccgcagc
gtgcgcgtggtggatcatccggatctgcaggatggcccggataacctgtggaacaccgaa
attgtgcagcgcgaagtgctgagctatctgcatagcgtgaaagatattcgcaccgtgatt
acctttgatcatcgcggcgtgagcagccatgcgaaccatgtggcggtgtatgaaggcgtg
ctgctggcgaaaaaaaacctgccgccgggcattctgtttctgagcctgcatacccgcgat
ctgctggaaaaatatgtgggcattctgagcaccgtgggctataccgtgggcattcatcgc
tgcggcggccgccgcaaccatgtgattctgattccgccgaccagcctgtttaccagcttt
agcgcgatgcgcaaacataaaacccagctggtgtggtttcgctatctgtttgtgtggttt
agcagctatagctatgtgaacgaagtgaaagaactgggcgtggcg
GC% Content for gene: 55.03%
CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): N/a
GC% Content for gene (codon optimized): N/a
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):